Spinal Afferents Research Articles

Neurological and cellular regulation of visceral hypersensitivity induced by chronic stress and colonic inflammation in rats

The role of inflammation in inducing visceral hypersensitivity (VHS) in ulcerative colitis patients remains unknown. We tested the hypothesis that acute ulcerative colitis-like inflammation does not induce VHS. However, it sets up molecular conditions such that chronic stress following inflammation exaggerates single-unit afferent discharges to colorectal distension. We used dextran sodium sulfate (DSS) to induce ulcerative colitis-like inflammation and a 9-day heterotypic chronic stress protocol in rats. DSS upregulated Nav1.8 mRNA in colon-responsive dorsal root ganglion (DRG) neurons, TRPV1 in colonic muscularis externae (ME) and BDNF in spinal cord without affecting the spike frequency in spinal afferents or VMR to CRD. By contrast, chronic stress did not induce inflammation but it downregulated Kv1.1 and Kv1.4 mRNA in DRG neurons, and upregulated TRPA1 and nerve growth factor in ME, which mediated the increase of spike frequency and VMR to CRD. Chronic stress following inflammation exacerbated spike frequency in spinal afferent neurons. TRPA1 antagonist suppressed the sensitization of afferent neurons. DSS-inflammation did not affect the composition or excitation thresholds of low-threshold and high-threshold fibers. Chronic stress following inflammation increased the percent composition of high-threshold fibers and lowered the excitation threshold of both types of fibers. We conclude that not all types of inflammation induce VHS, whereas chronic stress induces VHS in the absence of inflammation.

Functional role of peripheral opioid receptors in the regulation of cardiac spinal afferent nerve activity during myocardial ischemia

Thinly myelinated Aδ-fiber and unmyelinated C-fiber cardiac sympathetic (spinal) sensory nerve fibers are activated during myocardial ischemia to transmit the sensation of angina pectoris. Although recent observations showed that myocardial ischemia increases the concentrations of opioid peptides and that the stimulation of peripheral opioid receptors inhibits chemically induced visceral and somatic nociception, the role of opioids in cardiac spinal afferent signaling during myocardial ischemia has not been studied. The present study tested the hypothesis that peripheral opioid receptors modulate cardiac spinal afferent nerve activity during myocardial ischemia by suppressing the responses of cardiac afferent nerve to ischemic mediators like bradykinin and extracellular ATP. The nerve activity of single unit cardiac afferents was recorded from the left sympathetic chain (T₂-T₅) in anesthetized cats. Forty-three ischemically sensitive afferent nerves (conduction velocity: 0.32-3.90 m/s) with receptive fields in the left and right ventricles were identified. The responses of these afferent nerves to repeat ischemia or ischemic mediators were further studied in the following protocols. First, epicardial administration of naloxone (8 μmol), a nonselective opioid receptor antagonist, enhanced the responses of eight cardiac afferent nerves to recurrent myocardial ischemia by 62%, whereas epicardial application of vehicle (PBS) did not alter the responses of seven other cardiac afferent nerves to ischemia. Second, naloxone applied to the epicardial surface facilitated the responses of seven cardiac afferent nerves to epicardial ATP by 76%. Third, administration of naloxone enhanced the responses of seven other afferent nerves to bradykinin by 85%. In contrast, in the absence of naloxone, cardiac afferent nerves consistently responded to repeated application of ATP (n = 7) or bradykinin (n = 7). These data suggest that peripheral opioid peptides suppress the responses of cardiac sympathetic afferent nerves to myocardial ischemia and ischemic mediators like ATP and bradykinin.

Pancreatic Exocrine Insufficiency (PEI) in chronic pancreatitis (CP) and cystic fibrosis (CF) patients: Combining clinician insights and existing literature to develop a conceptual model to aid in the identification and management of PEI

Pancreatic Exocrine Insufficiency (PEI) in chronic pancreatitis (CP) and cystic fibrosis (CF) patients: Combining clinician insights and existing literature to develop a conceptual model to aid in the identification and management of PEI

Age-related changes of neurochemically different subpopulations of cardiac spinal afferent neurons in rats

This study investigated the effect of aging on cardiac spinal afferent neurons in the rat. A patch loaded with retrograde tracer Fast Blue (FB) was applied to all chambers of the rat heart. Morphological and neurochemical characteristics of labeled cardiac spinal afferent neurons were assessed in young (2months) and old (2years) rats using markers for likely unmyelinated (isolectin B4; IB4) and myelinated (neurofilament 200; N52) neurons.The number of cardiac spinal afferent neurons decreased in senescence to 15% of that found in young rats (1604 vs. 248). The size of neuronal soma as well as proportion of IB4+ neurons increased significantly, whereas the proportion of N52+ neurons decreased significantly in senescence. Unlike somatic spinal afferents, neurochemically different populations of cardiac spinal afferent neurons experience morphological and neurochemical changes related to aging. A major decrease in total number of cardiac spinal afferent neurons occurs in senescence. The proportion of N52+ neurons decreased in senescence, but it seems that nociceptive innervation is preserved due to increased proportion and size of IB4+ unmyelinated neurons.

Leptin-sensitive sensory nerves innervate white fat

Leptin, the primary white adipose tissue (WAT) adipokine, is thought to convey lipid reserve information to the brain via the circulation. Because WAT responds to environmental/internal signals in a fat pad-specific (FPS) manner, systemic signals such as leptin would fail to communicate such distinctive information. Saturation of brain leptin transport systems also would fail to convey increased lipid levels beyond that point. WAT possesses sensory innervation exemplified by proven sensory-associated peptides in nerves within the tissue and by viral sensory nerve-specific transneuronal tract tracer, H129 strain of herpes simplex virus 1 labeling of dorsal root ganglia (DRG) pseudounipolar neurons, spinal cord and central sensory circuits. Leptin as a paracrine factor activating WAT sensory innervation could supply the brain with FPS information. Therefore, we tested for and found the presence of the long form of the leptin receptor (Ob-Rb) on DRG pseudounipolar neurons immunohistochemically labeled after injections of Fluorogold, a retrograde tract tracer, into inguinal WAT (IWAT). Intra-IWAT leptin injections (300 ng) significantly elevated IWAT nerve spike rate within 5 min and persisted for at least 30 min. Intra-IWAT leptin injections also induced significant c-Fos immunoreactivity (ir), indicating neural activation across DRG pseudounipolar sensory neurons labeled with Fluorogold IWAT injections. Intraperitoneal leptin injection did not increase c-Fos-ir in DRG or the arcuate nucleus, nor did it increase arcuate signal transducer and activator of transcription 3 phosphorylation-ir. Collectively, these results strongly suggest that endogenous leptin secreted from white adipocytes functions as a paracrine factor to activate spinal sensory nerves innervating the tissue.

Opioids modulate cardiac spinal afferent responses to ischemia and other mediators

Cardiac spinal afferents are activated during myocardial ischemia. Although recent observations show that myocardial ischemia increases the concentrations of opioid peptides and that stimulation of peripheral opioid receptors inhibits chemical‐induced visceral and somatic nociception, the role of opioids in afferent signaling duirng pathophysiological events is unclear. Thus, we hypothesized that peripheral opioids modulate cardiac spinal afferent activity during myocardial ischemia. Nerve activity of single‐unit cardiac afferents in anesthetized cats was recorded from the left sympathetic chain (T2 – T5). Thirty‐three ischemically sensitive afferents (CV=0.34–3.90 m/s) with receptive fields in the left ventricle were identified by their responses to five min of regional ischemia. The responses of these afferents to repeat ischemia or ischemic mediators were further studied. First, epicardial application of naloxone (80 μmol), a non‐selective opioid receptor antagonist, enhanced the responses of 7 cardiac afferents to recurrent myocardial ischemia by 35%, whereas epicardial application of saline did not alter responses of 6 other afferents to repeat ischemia. Second, epicardial naloxone respectively facilitated the responses of other ischemically sensitive cardiac afferents to ATP (n=5) by 54% and bradykinin (BK, n=5) by 52%. In contrast, in the absence of naloxone, the afferents consistently responded to repeated application of ATP (n=5) or BK (n=5). These data suggest that peripheral opioid peptides modulate cardiac spinal afferents responses to ischemia and ischemic mediators like ATP and bradykinin. (Supported by NIH HL66217)

Mast cell expression of the serotonin1Areceptor in guinea pig and human intestine

Serotonin [5-hydroxytryptamine (5-HT)] is released from enterochromaffin cells in the mucosa of the small intestine. We tested a hypothesis that elevation of 5-HT in the environment of enteric mast cells might degranulate the mast cells and release mediators that become paracrine signals to the enteric nervous system, spinal afferents, and secretory glands. Western blotting, immunofluorescence, ELISA, and pharmacological analysis were used to study expression of 5-HT receptors by mast cells in the small intestine and action of 5-HT to degranulate the mast cells and release histamine in guinea pig small intestine and segments of human jejunum discarded during Roux-en-Y gastric bypass surgeries. Mast cells in human and guinea pig preparations expressed the 5-HT1A receptor. ELISA detected spontaneous release of histamine in guinea pig and human preparations. The selective 5-HT1A receptor agonist 8-hydroxy-PIPAT evoked release of histamine. A selective 5-HT1A receptor antagonist, WAY-100135, suppressed stimulation of histamine release by 5-HT or 8-hydroxy-PIPAT. Mast cell-stabilizing drugs, doxantrazole and cromolyn sodium, suppressed the release of histamine evoked by 5-HT or 8-hydroxy-PIPAT in guinea pig and human preparations. Our results support the hypothesis that serotonergic degranulation of enteric mast cells and release of preformed mediators, including histamine, are mediated by the 5-HT1A serotonergic receptor. Association of 5-HT with the pathophysiology of functional gastrointestinal disorders (e.g., irritable bowel syndrome) underlies a question of whether selective 5-HT1A receptor antagonists might have therapeutic application in disorders of this nature.

Expression of activating transcription factor 3 (ATF3) in uninjured dorsal root ganglion neurons in a lower trunk avulsion pain model in rats

Clinically, neuropathic pain is frequent and intense following brachial plexus injury. It is thought that brachial plexus pain is not generated by avulsed roots, but rather by non-avulsed roots, since the avulsed root could not possibly transmit action potentials to central nerves. The aim of this study was to evaluate pain behavior and activation of sensory neurons in a brachial plexus avulsion (BPA) model in rats. Fifteen male Wistar rats were used. In the BPA group, the C8-T1 roots were avulsed from the spinal cord with forceps at the lower trunk level (n = 5). In the naïve group, rats did not receive any procedures (n = 5). In the sham-operated group, the lower trunk was simply exposed (n = 5). Mechanical hyperalgesia of forelimbs corresponding to C6 and C7 dermatomes was measured using von Frey filaments every third day for 3 weeks. Activation of DRG neurons was immunohistochemically examined using anti-ATF3 (a marker for neuron activation) antibodies 21 days after surgery. Von Frey and immunohistochemical data between groups were analyzed using a Kruskal-Wallis test, followed by Mann-Whitney U tests. Bonferroni corrections were performed. Animals in the BPA group displayed significant mechanical hyperalgesia at the dermatome innervated by uninjured nerves continuing through day 21 compared with animals in the sham-operated group. ATF3-immunoreactive small and large DRG neurons were significantly activated in the BPA group (10.6 ± 9.5 and 5.2 ± 4.1 %, 39.7 ± 6.7 and 25.2 ± 10.3 %, 78.0 ± 9.1 and 53.7 ± 29.3 %) compared with the sham-operated group (0.7 ± 0.9 and 0 ± 0 %, 2.8 ± 2.0 and 1.0 ± 2.0 %, 3.9 ± 2.7 and 8.6 ± 10.1 %) at every level of C5, 6, and 7. In the naïve group, no DRG neurons were activated. ATF3-immunoreactive small and large DRG neurons were significantly activated at the level of C7 compared with C6 and C5, and significantly activated at the level of C6 compared with C5 in the BPA group. Expression of ATF3 in uninjured DRG neurons may contribute to pain following brachial plexus avulsion injury. Consequently, spared spinal sensory nerves may represent therapeutic targets for treatment of this pain.

Imaging stretch-activated firing of spinal afferent nerve endings in mouse colon

Spinal afferent neurons play a major role in detecting noxious and innocuous stimuli from visceral organs, such as the gastrointestinal tract. However, all our understanding about spinal afferents has been obtained from recordings of spinal afferent axons, or cell bodies that lie outside the gut wall, or peripheral organ they innervate. No recordings have been made directly from spinal afferent nerve endings, which is where sensory transduction occurs. We developed a preparation whereby recordings could be made from rectal afferent nerve endings in the colon, to characterize mechanisms underlying sensory transduction. Dorsal root ganglia (L6-S2) were removed from mice, whilst retaining neural continuity with the colon. Fluo-4-AM was used to record from rectal afferent nerve endings in myenteric ganglia and circular muscle at 36°C. In slack (unstretched) preparations of colon, no calcium transients were recorded from spinal afferent endings. However, in response to a maintained increase in circumferential diameter, a maintained discharge of calcium transients occurred simultaneously in multiple discrete varicosities along single axons of rectal afferents in myenteric ganglia and circular muscle. Stretch-activated calcium transients were resistant to hexamethonium and nifedipine, but were abolished by tetrodotoxin, CPA, BAPTA-AM, cobalt, gadolinium, or replacement of extracellular Na+ with NMDG. In summary, we present a novel preparation in which stretch-activated firing of spinal afferent nerve endings can be recorded, using calcium imaging. We show that circumferential stretch of the colon activates a maintained discharge of calcium transients simultaneously in varicosities along single rectal afferent endings in myenteric ganglia and circular muscle. Non-selective cation channels, TTX-sensitive Na+ channels and both extracellular calcium influx and intracellular Ca2+ stores are required for stretch-activated calcium transients in rectal afferent endings.

Challenges and opportunities of sensory plasticity after SCI

Even in cases of spinal cord injury (SCI) where sensory perceptions do not arise from stimuli applied to below-level regions, sensory input to the spinal cord, carried by spinal sensory afferents, still occurs and influences the central and autonomic nervous systems (CNS, ANS). This is true also of the vagal system which provides non-spinal innervation of viscera below many spinal cord injuries. It is therefore important to consider (1) how the neurochemical, anatomical, and electrophysiological properties of these sensory neurons, and the processing of the inputs by the CNS and ANS, is altered by SCI, (2) whether and how they may play a role in pathologies, and (3) how they may interact with treatment strategies. This Research Topic addresses plasticity of sensory systems after SCI, with a non-exclusive focus on systems below the level of the injury. POST-SCI AUTONOMIC DYSFUNCTIONS The ANS controls systems below the level of consciousness and this is often taken for granted until something goes awry. Those living with SCI are acutely aware of the functions regulated, or more often dysregulated, by the ANS. One of the most pressing of these issues is autonomic dysreflexia (AD), a chronic and com

Gastric Electrical Stimulation Decreases Gastric Distension-Induced Central Nociception Response through Direct Action on Primary Afferents

Background & AimsGastric electrical stimulation (GES) is an effective therapy to treat patients with chronic dyspepsia refractory to medical management. However, its mechanisms of action remain poorly understood.MethodsGastric pain was induced by performing gastric distension (GD) in anesthetized rats. Pain response was monitored by measuring the pseudo-affective reflex (e.g., blood pressure variation), while neuronal activation was determined using c-fos immunochemistry in the central nervous system. Involvement of primary afferents was assessed by measuring phosphorylation of ERK1/2 in dorsal root ganglia.ResultsGES decreased blood pressure variation induced by GD, and prevented GD-induced neuronal activation in the dorsal horn of the spinal cord (T9–T10), the nucleus of the solitary tract and in CRF neurons of the hypothalamic paraventricular nucleus. This effect remained unaltered within the spinal cord when sectioning the medulla at the T5 level. Furthermore, GES prevented GD-induced phosphorylation of ERK1/2 in dorsal root ganglia.ConclusionsGES decreases GD-induced pain and/or discomfort likely through a direct modulation of gastric spinal afferents reducing central processing of visceral nociception.

Visualization of spinal afferent innervation in the mouse colon by AAV8‐mediated GFP expression

Primary afferent neurons whose cell bodies reside in thoracolumbar and lumbosacral dorsal root ganglia (DRG) innervate colon and transmit sensory signals from colon to spinal cord under normal conditions and conditions of visceral hypersensitivity. Histologically, these extrinsic afferents cannot be differentiated from intrinsic fibers of enteric neurons because all known markers label neurons of both populations. Adeno-associated virus (AAV) vectors are capable of transducing DRG neurons after intrathecal administration. We hypothesized that AAV-driven overexpression of green fluorescent protein (GFP) in DRG would enable visualization of extrinsic spinal afferents in colon separately from enteric neurons. Recombinant AAV serotype 8 (rAAV8) vector carrying the GFP gene was delivered via direct lumbar puncture. Green fluorescent protein labeling in DRG and colon was examined using immunohistochemistry. Analysis of colon from rAAV8-GFP-treated mice demonstrated GFP-immunoreactivity (GFP-ir) within mesenteric nerves, smooth muscle layers, myenteric plexus, submucosa, and mucosa, but not in cell bodies of enteric neurons. Notably, GFP-ir colocalized with CGRP and TRPV1 in mucosa, myenteric plexus, and globular-like clusters surrounding nuclei within myenteric ganglia. In addition, GFP-positive fibers were observed in close association with blood vessels of mucosa and submucosa. Analysis of GFP-ir in thoracolumbar and lumbosacral DRG revealed that levels of expression in colon and L6 DRG appeared to be related. These results demonstrate the feasibility of gene transfer to mouse colonic spinal sensory neurons using intrathecal delivery of AAV vectors and the utility of this approach for histological analysis of spinal afferent nerve fibers within colon.

Firing patterns and functional roles of different classes of spinal afferents in rectal nerves during colonic migrating motor complexes in mouse colon

The functional role of the different classes of visceral afferents that innervate the large intestine is poorly understood. Recent evidence suggests that low-threshold, wide-dynamic-range rectal afferents play an important role in the detection and transmission of visceral pain induced by noxious colorectal distension in mice. However, it is not clear which classes of spinal afferents are activated during naturally occurring colonic motor patterns or during intense contractions of the gut smooth muscle. We developed an in vitro colorectum preparation to test how the major classes of rectal afferents are activated during spontaneous colonic migrating motor complex (CMMC) or pharmacologically induced contraction. During CMMCs, circular muscle contractions increased firing in low-threshold, wide-dynamic-range muscular afferents and muscular-mucosal afferents, which generated a mean firing rate of 1.53 ± 0.23 Hz (n = 8) under isotonic conditions and 2.52 ± 0.36 Hz (n = 17) under isometric conditions. These low-threshold rectal afferents were reliably activated by low levels of circumferential stretch induced by increases in length (1-2 mm) or load (1-3 g). In a small proportion of cases (5 of 34 units), some low-threshold muscular and muscular-mucosal afferents decreased their firing rate during the peak of the CMMC contractions. High-threshold afferents were never activated during spontaneous CMMC contractions or tonic contractions induced by bethanechol (100 μM). High-threshold rectal afferents were only activated by intense levels of circumferential stretch (10-20 g). These results show that, in the rectal nerves of mice, low-threshold, wide-dynamic-range muscular and muscular-mucosal afferents are excited during contraction of the circular muscle that occurs during spontaneous CMMCs. No activation of high-threshold rectal afferents was detected during CMMCs or intense contractile activity in naïve mouse colorectum.

Opioid receptors modulate excitatory cardiovascular reflex responses to myocardial ATP stimulation through a P2 receptor mechanism

Activation of cardiac spinal afferents evokes excitatory cardiovascular reflexes. We have demonstrated that extracellular ATP stimulates ischemia‐sensitive cardiac afferents through P2 receptor mechanisms. The present study tested the hypothesis that through activation of P2 receptors ATP provokes excitatory cardiovascular reflexes and the reflexes are facilitated by blockade of opioid receptors. Hemodynamic parameters were recorded in anesthetized rats after bilateral vagotomy. Intrapericardial injection of α,β‐meATP (31–500 nmol), a P2X receptor agonist, increased mean arterial pressure (MAP) by 4±2 to 49±4 mmHg in a dose‐dependent manner in seven rats. After blockade of cardiac neural transmission with intrapericardial procaine, the α,β‐meATPevoked pressor responses were almost eliminated (n=6). Application of α,β‐meATP‐evoked consistent reflexes in six control rats. Intrapericardial ADP, a P2Y receptor agonist, also increased MAP in five rats by 9 to 13±3 mmHg, although we observed no changes in two and a decrease in two rats. Finally, blockade of opioid receptors with naloxone enhanced the α,β‐meATP‐mediated reflexes from increase in MAP by 27±3 to 37±3 mmHg (n=6). These data suggest that extracellular ATP provokes cardiac sympathetic afferent reflexes mainly through P2X receptor mechanisms and that the magnitude of these reflexes is partially dependent on intact opioid receptors.

A Novel pH Conditioned Cl‐ Conductance in Nodose Ganglia Neurons

pH sensitivity has been studied extensively in spinal afferents of dorsal root ganglia neurons but rarely in vagal afferents of nodose ganglia (NG) neurons. Here, we report a large and prolonged inward current evoked following 2 or 3 brief (10s) exposures to low pH (6.0) in 16 of 22 (70%) isolated nodose neurons using whole‐cell patch‐clamp technique. This pH‐conditioned current (pH‐I) reached its peak of 904.3±159.9pA (n=15) at ~5 minutes and returned to baseline within 10–15 minutes. Replacing Ca2+ and Na+ in the extracellular solution with NMDG or intracellular K+ with Cs+ did not significantly decrease the current. However pH‐I was eliminated by using a low Cl− (4mM) pipette solution. The I‐V plot showed a reversal potential of ~0mV with equivalent intra‐and extracellular [Cl]− (133mM) in the absence of permeable cations. Replacing [Cl]o− with aspartate shifted the reversal potential to a more positive voltage. pH‐I was reduced by tamoxifen, an inhibitor of “swell‐activated Cl− current” (SAC), but not by niflumic acid (blocker of the Ca2+ activated Cl− channel). Adding hypoosmotic solution, which activates SAC, did not further increase peak pH‐I. Conversely evoking the pH‐I did not increase the SAC current. Opening of this pH conditioned Cl− conductance may contribute to sustained depolarization of vagal afferents and potentiate a beneficial reflex sympathoinhibition during myocardial and gastrointestinal ischemia/acidosis (NIH‐HL14388).

Characterization of thromboxane A2 receptor and TRPV1 mRNA expression in cultured sensory neurons

Thromboxane A2 (TxA2) is an arachidonic acid metabolite that stimulates platelet aggregation and vasoconstriction when released from platelets and other cell types during tissue trauma. More recent research has demonstrated that TxA2 can also stimulate vagal and spinal sensory nerves. The purpose of this study was twofold. One, we compared the expression of the TxA2 receptor (TxA2R) in neurons from two sensory ganglia: the nodose ganglion (NG) containing cell bodies of vagal afferent nerves and the thoracic dorsal root ganglion (DRG) containing cell bodies of spinal afferent nerves. Two, we determined if TxA2R co-localizes with mRNA for the nociceptive marker, TRPV1, which is the receptor for the noxious substance capsaicin. We found a greater percentage of neurons in the NG that are positive for TxA2R expression than in the DRG. We also found that there was no correlation of expression of TxA2R with TRPV1. These data suggest that while TxA2R is expressed in both vagal and spinal neurons, TxA2 may elicit stronger vagal or parasympathetic reflexes in the rabbit when released during tissue trauma depending on the location of release. Our data also indicate that TxA2 is likely to stimulate both nociceptive and non-nociceptive neurons thereby broadening the types of neurons and reflexes that it may excite.

On the concept of spinocerebellum

From an evolutionary point of view, the cerebellum consists of three regions: the archicerebellum, which is dominant in aquatic animals; the paleocerebellum, which developed during terrestrial locomotion in parallel with the spinal cord; and the neocerebellum, which dominates in mammals and developed along with the cerebral hemispheres. The paleocerebellum, identified mainly with the vermis, is often called the spinocerebellum, although spinal afferents project only to the vermal area of the anterior lobe and to a subregion of the posterior lobe. Therefore, the name spinocerebellum should not be synonymous with vermis and should not imply a lack of cortical afferents. The article by Coffman et al. (1) demonstrated, by means of retrograde viral tracing, that the cerebellar vermis—predominantly the posterior region—receives projections from the motor areas in the cerebral cortex. The authors concluded that their results challenge the classical view on the vermis and indicate that this structure should no longer be considered as entirely isolated from the cerebral cortex. In the interest of the scientific debate regarding how afferent information is organized over the cerebellar cortex, we would like to mention that (i) projections from the motor cortex to the vermal area of the anterior lobe and (ii) their complex distribution have been described in an article published in 1968 (2). In this study we investigated the spinal and cortical projections to the cat anterior lobe both in the vermis and in the pars intermedia along lobuli I–V. We analyzed electrophysiologically the projections mediated by the mossy fiber (MF) and the climbing fiber (CF) pathways from the forelimb (F-CORT) and the hindlimb (H-CORT) areas of the sensorimotor cortex and made a comparison with the known projections from forelimb and hindlimb peripheral nerves. Stimulation of F-CORT and H-CORT evokes in the anterior lobe an early response at a latency of 3–3.5 ms due to the MF input and a later response at a latency of 13–16 ms due to the CF input.

Changes in microglial activation within the hindbrain, nodose ganglia, and the spinal cord following subdiaphragmatic vagotomy

Damage to peripheral nerve branches triggers activation of microglia in CNS areas containing motor neuron soma and primary afferent terminals of the damaged fibers. Furthermore, microglial activation occurs in areas containing the soma and terminals of spared nerve branches of a damaged nerve. Because the abdominal viscera are innervated by spinal afferents as well as vagal afferents and efferents, we speculated that spinal nerves might respond like spared nerve branches following damage to vagal fibers. Therefore, we tested the hypothesis that damage to the abdominal vagus would result in microglial activation in vagal structures—the nucleus of the solitary tract (NTS), dorsal motor nucleus of the vagus nerve (DMV), and nodose ganglia (NG)—as well as spinal cord (SC) segments that innervate the abdominal viscera. To test this hypothesis, rats underwent subdiaphragmatic vagotomy or sham surgery and were treated with saline or the microglial inhibitor, minocycline. Microglial activation was determined by quantifying changes in the intensity of fluorescent staining with a primary antibody against ionizing calcium adapter binding molecule 1 (Iba1). We found that subdiaphragmatic vagotomy significantly activated microglia in the NTS, DMV, and NG two weeks post-vagotomy. Microglial activation remained significantly increased in the NG and DMV for at least 42 days. Surprisingly, vagotomy significantly decreased microglial activation in the SC. Minocycline treatment attenuated microglial activation in all studied areas. Our results indicate that microglial activation in vagal structures following abdominal vagal damage is accompanied by suppression of microglial activation in associated areas of the spinal cord.

Hyperpolarization-activated cyclic nucleotide-gated cation channel subtypes differentially modulate the excitability of murine small intestinal afferents

To assess the role of hyperpolarization-activated cyclic nucleotide-gated cation (HCN) channels in regulating the excitability of vagal and spinal gut afferents. The mechanosensory response of mesenteric afferent activity was measured in an ex vivo murine jejunum preparation. HCN channel activity was recorded through voltage and current clamp in acutely dissociated dorsal root ganglia (DRG) and nodose ganglia (NG) neurons retrogradely labeled from the small intestine through injection of a fluorescent marker (DiI). The isoforms of HCN channels expressed in DRG and NG neurons were examined by immunohistochemistry. Ramp distension of the small intestine evoked biphasic increases in the afferent nerve activity, reflecting the activation of low- and high-threshold fibers. HCN blocker CsCl (5 mmol/L) preferentially inhibited the responses of low-threshold fibers to distension and showed no significant effects on the high-threshold responses. The effect of CsCl was mimicked by the more selective HCN blocker ZD7288 (10 μmol/L). In 71.4% of DiI labeled DRG neurons (n = 20) and 90.9% of DiI labeled NG neurons (n = 10), an inward current (I(h) current) was evoked by hyperpolarization pulses which was fully eliminated by extracellular CsCl. In neurons expressing I(h) current, a typical "sag" was observed upon injection of hyperpolarizing current pulses in current-clamp recordings. CsCl abolished the sag entirely. In some DiI labeled DRG neurons, the I(h) current was potentiated by 8-Br-cAMP, which had no effect on the I(h) current of DiI labeled NG neurons. Immunohistochemistry revealed differential expression of HCN isoforms in vagal and spinal afferents, and HCN(2) and HCN(3) seemed to be the dominant isoform in DRG and NG, respectively. HCNs differentially regulate the excitability of vagal and spinal afferent of murine small intestine.

Liver Afferents Contribute to Water Drinking-Induced Sympathetic Activation in Human Subjects: A Clinical Trial

Water drinking acutely increases sympathetic activity in human subjects. In animals, the response appears to be mediated through transient receptor potential channel TRPV4 activation on osmosensitive hepatic spinal afferents, described as osmopressor response. We hypothesized that hepatic denervation attenuates water drinking-induced sympathetic activation. We studied 20 liver transplant recipients (44±2.6 years, 1.2±0.1 years post transplant) as model of hepatic denervation and 20 kidney transplant recipients (43±2.6 years, 0.8±0.1 years post transplant) as immunosuppressive drug matched control group. Before and after 500 ml water ingestion, we obtained venous blood samples for catecholamine analysis. We also monitored brachial and finger blood pressure, ECG, and thoracic bioimpedance. Plasma norepinephrine concentration had changed by 0.01±0.07 nmol/l in liver and by 0.21±0.07 nmol/l in kidney transplant recipients (p<0.05 between groups) after 30–40 minutes of water drinking. While blood pressure and systemic vascular resistance increased in both groups, the responses tended to be attenuated in liver transplant recipients. Our findings support the idea that osmosensitive hepatic afferents are involved in water drinking-induced sympathetic activation in human subjects.Trial RegistrationClinicalTrials.gov NCT01237431